Vincent Van Deuren

Synthetic fluorophores for live-cell fluorescence microscopy and biosensing - Nature Chemical Biology Performing live-cell microscopy experiments with high spatial and temporal resolution requires fluorophores with highly optimized properties. This Review examines the progress in developing synthetic ...

🚨Curious how synthetic fluorophores simplify microscopy? Our review covers applications such as super-resolution microscopy and biosensing. 🎉👩‍🔬Congrats to my first PhD student, Agnese Nicoli, for the fearless deep dive! 🔬 ⚗️
D-CHAB @ethz.ch
www.nature.com/articles/s41...

NanoBlocks: creating fluorescent biosensors from affinity binders using competitive binding Genetically-encoded fluorescent biosensors have revolutionized our understanding of complex systems by permitting the in situ observation of chemical activities. However, only a comparatively small se...

New preprint! Our work together with @agtebo.bsky.social : our 'NanoBlocks' strategy to generate fluorescent biosensors from binders such as nanobodies. Proof-of-concept sensors with ALFA nanobody & Erbin PDZ domain shows the concept in vitro, in cells, and for FACS.
www.biorxiv.org/cgi/content/...

www.biorxiv.org/content/10.1...

🚨New preprint 🚨Our 'NanoBlocks' strategy to generate fluorescent biosensors from binders such as nanobodies. Proof-of-concept sensors with ALFA nanobody & Erbin PDZ domain shows the concept in vitro, in cells, and for FACS. Have an antibody-like binder? Turn it into a biosensor using NanoBlocks.

PS. mStayGold as a FRET donor makes for highly photostable genetically-encoded biosensors! Definitely recommended if you want to do high-intensity/prolonged activity imaging.

@toonvanthillo.bsky.social @robinvde.bsky.social @dedeckerlab.bsky.social

Reduced Artifacts in Nanoscale Ratiometric Imaging and Sensing via Point-Spread Function Matching Ratiometric analysis of two or more fluorescence signals is a staple of quantitative imaging. However, this analysis becomes complicated at (near-) diffraction limited resolutions due to differences in how the emission colors are imaged by the microscope optics. We investigate this and find that point-spread function (PSF) mismatch between different emission wavelengths readily introduces spurious structuring in ratiometric images. We develop a postprocessing strategy that can correct for this effect by matching the respective PSFs, thus eliminating this deviation. To demonstrate this approach in live cells, we develop a highly photostable FRET biosensor against protein kinase A (PKA) activity and apply it to diffraction-limited biosensing, demonstrating that the spurious structuring can indeed be removed from the analyzed images. Our work further supports the feasibility of high-resolution ratiometric imaging using stable molecular probes and suitable correction strategies.

Luckily, we describe a straightforward post-processing strategy to correct for this effect. More information inside!
pubs.acs.org/doi/10.1021/...

Do you want to do nanoscale ratiometric fluorescence imaging? Make sure to account for point-spread-function (PSF) mismatch between emission channels! The differences in image formation causes the appearance of spurious structuring, resulting in misleading ratiometric images

🚨 Want a fast confocal? Struggling to capture dynamics in 3D? Lacking a system capable of 3D DNA-PAINT imaging 🧬✨?

TriScan – a single-molecule sensitive line-scan confocal with the speed of a widefield!

Preprint: biorxiv.org/content/10.1101/2023.04.11.536163v2

#Microscopy #DNApaint #Confocal